The long term objectives of this subproject are to understand the structure and function of gonococcal iron regulated membrane proteins. Utilizing a variety of genetic, biochemical, immunological, and molecular biological techniques, we will try to understand how gonococci acquire iron from the human proteins transferrin and lactoferrin. Specific emphasis will be on proteins involved in the transferrin receptor, which includes a 100 Kd transferrin binding protein and probably a second unrelated protein of 85 Kd. Genes for these proteins will be cloned, sequenced, and mutagenized by insertion of a transposon, and will be reintroduced into wild-type gonococci to make "molecular knockout mutants" for each protein. These molecular mutants will be studied for alterations in transferrin binding and iron utilization, and also for effects on pathogenicity in an endometrial organ culture model. Similar studies will focus on a 105 Kd lactoferrin binding protein which apparently is the lactoferrin receptor. Other experiments will examine the mode of regulation of these surface proteins. Antibodies will be raised against these proteins and will be studied for their potentially protective effects (decreased Fe uptake; bactericidal and/or used in project 3 for study of the effects of alterations in iron transport on urethral pathogenicity in human volunteers.